Effective isolation of nucleic acids from biological samples (e.g. cultured cells, tissue, viruses) is an essential prerequisite for efficient downstream amplification, detection, and quantification of specific genetic sequences via quantitative polymerase chain reaction (qPCR). The extraction process requires lysing the cells with harsh extraction reagents, such as detergents or enzymes, thereby resulting in a mixture of nucleic acids, cellular debris and extraction reagents. The nucleic acids are then separated/purified from the cellular debris and extraction reagents using a variety of techniques (e.g. organic solvent extraction, chromatography, centrifugation, dialysis). These techniques can be very time-consuming, tedious, and often require multiple washing steps. By way of example, commercially-available nucleic acid isolation kits require approximately 15 minutes to over one hour to complete, largely due to the multiple washing steps required to sufficiently separate the nucleic acids from the cellular debris and extraction reagents. Consequently, it has been suggested that as much as 15% of all molecular biology research time is devoted to purification.
In view of the foregoing, various attempts have been made to reduce the time associated with isolating nucleic acids from a biological sample. By way of example, Kelso, United States Patent Application No. 20090246782 discloses a system, device, and method for performing biological reactions. More specifically, the system contemplates placing a sample in a first chamber. The first chamber includes first processing reagents to generate a processed sample. The processed sample is moved through a water and alcohol immiscible, hydrophobic, or lipophilic barrier to a second chamber. The processed sample is treated in said second chamber with second processing reagents to generate a further processed sample.
While functional for its intended purpose, the system disclosed in the '782 application has certain limitations. For example, the reagents and immiscible phase of the system disclosed in the '782 application must be confined within corresponding chambers. As a result, the system requires the use of an external pump or two-axis magnet to move the processed sample between the chambers. It can be appreciated that the use of an external pump may have undesired effects on the sample. Alternatively, the use of a two-axis magnet may add unwanted cost and complexity to the system. In addition, the use of a plurality of chambers to isolate the nucleic acids from a biological sample may limit the throughput of the system.
Therefore, it is a primary object and feature of the present invention to provide a device for and a method of extracting and purifying a fraction from cultured cells, tissue samples and other biological materials.
It is a further object and feature of the present invention to provide a device for and a method of extracting and purifying a fraction from cultured cells, tissue samples and other biological materials that is simpler and more efficient than prior devices and methods.
It is a still further object and feature of the present invention to provide a device for and a method of extracting and purifying a fraction from cultured cells, tissue samples and other biological materials that has higher throughput than prior devices and methods.
In accordance with the present invention, a device is provided for facilitating extraction of a fraction from a biological sample. The biological sample includes non-desired material and a fraction-bound solid phase substrate. The device includes an input zone for receiving the biological sample therein and a phase-gate zone for receiving an isolation buffer therein. An output zone receives a reagent therein. A force is movable between a first position adjacent the input zone and a second position adjacent the output zone. The force urges the fraction-bound solid phase substrate from the input zone, through the phase-gate zone and into the output zone.
It is contemplated for the isolation buffer to be an oil. The oil prevents the non-desired material from passing therethrough. The device further includes a first channel has an input in communicating with the input zone and an output communicating with the phase-gate zone. The input of the first channel is larger than the output of the first channel. The first channel is partial defined by first and second sidewalls. The first and second sidewalls converge from the input of the first channel to the output of the first channel. In addition, the device may include a second channel having an input in communicating with the phase-gate zone and an output communicating with the output zone. The input of the second channel is larger than the output of the second channel and the input of the first channel is larger than the output of the first channel. The second channel is partial defined by first and second sidewalls. The first and second sidewalls defining the second channel converge from the input of the second channel to the output of the second channel.
In accordance with a further aspect of the present invention, a device is provided for facilitating extraction of a fraction from a biological sample. The biological sample includes non-desired material and a fraction-bound solid phase substrate. The device includes an input zone for receiving the biological sample therein and an output zone for receiving a reagent therein. A pathway interconnects the input zone and the output zone. A force field is movable between a first position adjacent the input zone and a second position adjacent the output zone. The force urges the fraction-bound solid phase substrate from the input zone, through the pathway and into the output zone.
An isolation buffer is disposed in the pathway. The isolation buffer prevents the non-desired material from passing therethrough. The pathway includes a phase-gate zone for housing the isolation buffer. The pathway is at least partially defined by a first channel has an input in communicating with the input zone and an output communicating with the phase-gate zone. The input of the first channel is larger than the output of the first channel. The first channel is partially defined by first and second sidewalls. The first and second sidewalls converge from the input of the first channel to the output of the first channel. The pathway may be further defined by a second channel having an input in communicating with the phase-gate zone and an output communicating with the output zone. The input of the second channel is larger than the output of the second channel. The second channel is partially defined by first and second sidewalls. The first and second sidewalls converge from the input of the second channel to the output of the second channel.
In accordance with a still further aspect of the present invention, a method is provided for facilitating extraction of a fraction from a biological sample. The biological sample includes non-desired material and a fraction-bound solid phase substrate. The method includes the steps of depositing the biological sample in an input zone and drawing the fraction-bound phase substrate from the input zone, through a phase-gate zone and into the output zone with a force.
The phase-gate zone includes an isolation buffer that prevents the non-desired material from passing therethrough. The isolation buffer may be an oil. The method may include the additional step of interconnecting the input zone and the phase-gate zone with a channel having an input in communication with the input zone and an output in communication with the phase-gate zone. The input of the channel is larger than the output of the channel. In addition, the phase-gate zone and the output zone may be interconnected with a channel having an input in communication with the phase-gate zone and an output in communication with the output zone. The input of the channel is larger than the output of the channel.